EP0325723A2 - Procédé pour la détermination de facteurs de coagulation du sang et réactif utilisé dans ce but - Google Patents

Procédé pour la détermination de facteurs de coagulation du sang et réactif utilisé dans ce but Download PDF

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Publication number
EP0325723A2
EP0325723A2 EP88119491A EP88119491A EP0325723A2 EP 0325723 A2 EP0325723 A2 EP 0325723A2 EP 88119491 A EP88119491 A EP 88119491A EP 88119491 A EP88119491 A EP 88119491A EP 0325723 A2 EP0325723 A2 EP 0325723A2
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EP
European Patent Office
Prior art keywords
blood clotting
clotting factor
monoclonal antibody
determination
reagent
Prior art date
Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
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Application number
EP88119491A
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German (de)
English (en)
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EP0325723A3 (fr
Inventor
Yasuo C/O Daiichi Pure Chemicals Co. Ltd. Sakai
Fujio C/O Daiichi Pure Chemicals Co. Ltd. Takei
Takashi C/O Daiichi Pure Chem. Co. Ltd Matsumoto
Makiko C/O Daiichi Pure Chemicals Co. Ltd. Maeda
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Daiichi Pure Chemicals Co Ltd
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Daiichi Pure Chemicals Co Ltd
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Publication date
Application filed by Daiichi Pure Chemicals Co Ltd filed Critical Daiichi Pure Chemicals Co Ltd
Publication of EP0325723A2 publication Critical patent/EP0325723A2/fr
Publication of EP0325723A3 publication Critical patent/EP0325723A3/fr
Withdrawn legal-status Critical Current

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    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
    • G01N33/48—Biological material, e.g. blood, urine; Haemocytometers
    • G01N33/50—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
    • G01N33/86—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing involving blood coagulating time or factors, or their receptors
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
    • G01N33/48—Biological material, e.g. blood, urine; Haemocytometers
    • G01N33/50—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
    • G01N33/53—Immunoassay; Biospecific binding assay; Materials therefor
    • G01N33/543—Immunoassay; Biospecific binding assay; Materials therefor with an insoluble carrier for immobilising immunochemicals
    • G01N33/54313—Immunoassay; Biospecific binding assay; Materials therefor with an insoluble carrier for immobilising immunochemicals the carrier being characterised by its particulate form
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N2333/00—Assays involving biological materials from specific organisms or of a specific nature
    • G01N2333/435—Assays involving biological materials from specific organisms or of a specific nature from animals; from humans
    • G01N2333/745—Assays involving non-enzymic blood coagulation factors
    • G01N2333/75—Fibrin; Fibrinogen

Definitions

  • This invention relates to a method and a reagent for detecting and quantitatively analyzing, with the use of monoclonal antibody, a blood clotting factor contained in a sample.
  • FDP cross-linked fibrinogen degradation product
  • Determination of FDP therefore, is very important for the detection of blood clotting or fibrinolysis in a living body, and knowing the result of that determination precisely and speedily is clinically imperative.
  • the latex agglutination test on a slide plate by the use of latex particles is judged with the naked eye. This inevitably leaves the judgment to the discretion of the person observing and renders the determination qualitative.
  • the enzymatic enzyme immunoassay for the quantitative analysis of FDP (Japanese Patent Laid-open No. 233,553/1985) takes considerable time and requires a certain technique for the determination.
  • the nephelometric method for quantitative determination with the use of a latex can be considered. This method, however, is liable to sustain a non-specific reaction.
  • the method usually relies upon a colorimetric technique for the determination, either use of a blank sample is necessary or changes in absorbance during the reaction must be detected. This requires a special measuring device for a precise determination.
  • the inventors have undertaken extensive studies in order to to overcome the drawbacks existing in conventional methods, and to develop a method for speedily determining a clotting factor.
  • the inventors have found that the use of a monoclonal antibody specific to a blood clotting factor and an insoluble carrier reagent to produce an agglutination, and the comparison of the amounts of non-­ agglutinated particles and agglutinated particles ensures an easy means for the quantitative determination of a blood clotting factor.
  • the finding has led to the completion of this invention.
  • an object of this invention is to provide a method for the determination of blood clotting factor comprising reacting a sample containing a blood clotting factor with an insoluble carrier reagent which is sensitized by a monoclonal antibody specific to the blood clotting factor, and comparing the amount of non-agglutinated particles with the amount of agglutinated particles produced.
  • Another object of the present invention is to provide a reagent for blood clotting factor determination comprising an insoluble carrier having a particle size of 0.5 to 2 ⁇ m which is sensitized with monoclonal antibody specific to the blood clotting factor.
  • Monoclonal antibodies used in the present invention can be prepared according to a conventional method. Beside FDP, protein C, ⁇ -TG, AT III, PLG, ⁇ 2P1, thrombin, FX III, or the like can be used as a blood clotting factor. These monoclonal antibodies can be used after they are selected by conventional method. It is preferable, however, for the purpose of avoiding interference related to complements, thus reducing the occurrence of non-specific reactions, to convert these into F(ab′)2 or Fab through digestion by the use of pepsin or papain. Given as examples of monoclonal antibodies having specificity for a blood clotting factor are the monoclonal antibody against FDP (Japanese Patent Laid-open No. 166,698/1985), and the like.
  • insoluble carrier particles used in this invention include polyamide-type, polyvinylidene chloride-­type, polyvinyl chloride-type, acryl-type, polyvinyl acetate-type, polystyrene-type, polypropylene-type, polyepoxy-type, urethane-type, polyethylene-type, agarose-­type chitosan-type, pullulan-type polymer particles as well as other types of polymer particles. It is desirable that these insoluble carrier particles contain a certain proportion of an acid group such as carboxyl group, sulfonic acid group, or the like.
  • a preferable method for preparing these types of polymers is to copolymerize a monomer having no acid group and a monomer having an acid group.
  • those having a degree of denaturing with an acid group in the range of 0.1 to 10% is preferably.
  • insoluble carrier particles which are commercially available are SFL-140L series products, SFL-­155L series products, SFL-200L series products, and SFL-140S series products, all manufactured by Japan Synthetic Rubber Co., Ltd.; N-series products manufactured by Sekisui Chemical Co., Ltd.; carboxy-denatured latex particles manufactured by Dow Chemical Co.; K-series latex particles manufactured by Rhone-Poulenc Co.; and the like.
  • these insoluble carrier particles those having a particle size preferably of 0.5 to 2 ⁇ m, and particularly of 0.6 to 1.2 ⁇ m, can be used.
  • Sensitization of insoluble carrier particles may be performed according to a conventional method. For instance, it can be performed by a physical adsorption method which comprises adding a monoclonal antibody buffer solution to a suspension of insoluble carrier particles, allowing the solution to stand for a duration of 1 hour to overnight, and eliminating the non-sensitized monoclonal antibody by a suitable means such as centrifugation.
  • a chemical covalent bonding method which comprises sensitizing an insoluble carrier with a monoclonal antibody using a known bi-crosslinking reagent or a bonding agent is desirable.
  • the generally applicable concentration of monoclonal antibody used to sensitize insoluble carrier particles is approximately 25 to 300 ⁇ g/ml and that of insoluble carrier particles is in the range of 0.2 to 2%. These concentrations, however, may be varied depending on the type and concentration of clotting agent to be measured, the type and characteristics of monoclonal antibody employed, as well as the property, particle size, and the degree of denaturing with an acid, of the insoluble carrier.
  • insoluble carrier sensitized with a monoclonal antibody is treated with bovine serum albumin (hereinafter abbreviated as "BSA”) or a surface active agent of various kinds as required, and is stored as a suspension in a buffer solution or as a lyophilized product.
  • BSA bovine serum albumin
  • Determination of a blood clotting factor according to the present invention is performed by adding a sample containing a blood clotting factor to be detected to an insolubilized antibody reagent, incubating the mixture, and measuring for comparison the amounts of non-agglutinated particles and agglutinated particles produced.
  • Plasma, serum, body fluid, or the like can be used as a sample. Incubation is carried out at a temperature of 20 to45°C for 3 to 30 minutes.
  • the amounts of non-agglutinated particles and agglutinated particles can be determined by counting, by means of a laser scattering method or the like, the number of particles having a particle size corresponding to the insolubilized antibody reagent and the number of particles having a particle size corresponding to the agglutinated particles produced. More specifically, a calibration curve is prepared in advance from the ratio (reaction ratio) of agglutinated particles to non-agglutinated particles produced using a clotting factor sample of known concentration. This calibration curve is then used as a standard for determining the amount of the clotting factor in a sample.
  • an orifice bore for apertures of 20 to 50 ⁇ m is suitable. Precise determination of clotting factors is possible using this type of particle measurement device by the application of the method of this invention.
  • Major advantages of this invention are: (i) it does not require the use of a defibrinogen, (ii) it can provide a highly sensitive measurement, (iii) the procedure is very simple requiring a very short time period for the measurement; less than 30 minutes starting from blood sampling, (iv) the method can exclude any non-specific reaction, e.g, it is free from the influences due to a rheumatoid factor.
  • the method provides a simple and speedy determination of clotting factors in living bodies, and is useful in a variety of the fields, including medical treatment and clinical inspection.
  • a 10% polystyrene latex suspension having a 1.0 ⁇ m particle size (SFL-140L manufactured by Japan Synthetic Rubber Co., Ltd.) was diluted to a 10-fold volume using a 0.05 M glycin buffer (pH 8.0).
  • the diluted suspension was mixed with an equivalent amount of a 100 ⁇ g/ml FDP monoclonal antibody (monoclonal antibody 03202 described in Japanese Patent Laid-open No.166,698/1985) and the mixture was allowed to stand for 2 hours at 4°C. Then, the same buffer solution containing 0.5% BSA in an amount of 1 part by volume per 100 parts by volume of the mixture was added.
  • An insolubilized FDP antibody reagent was prepared in the same manner as in Example 1 except that polystyrene latices having a 0.8 ⁇ m particle size and the monoclonal antibody 03204 described in Japanese Patent Laid-open No.166,698/1985 were used.
  • a calibration curve was prepared to determine the amount of FDP by mixing 50 ⁇ l of a standard FDP or 50 ⁇ l a sample with 50 ⁇ l of the insolubilized antibody reagent prepared in (1) above and following the same manner as in (2) (i) through (iii) of Example 1. The results are shown in Fig. 3.
  • a 10% polystyrene latex suspension having a 1.0 ⁇ m particle size (manufactured by Japan Synthetic Rubber Co., Ltd.) was diluted to a 10-fold volume using a 0.05 M glycin buffer (pH 8.0).
  • the diluted suspension was mixed with each 100 ⁇ g/ml ⁇ -TG monoclonal antibody, designated as 11211 and 11224. Each mixture was allowed to stand for 2 hours at an ambient temperature. Then, each was mixed with an equivalent amount of the same glycin buffer solution containing 0.5% BSA and allowed to stand for 2 hours at 4°C. The mixture was subjected to centrifugation to wash out and eliminate non-sensitized monoclonal antibodies.
  • the residue was ultimately made into a suspension containing 0.2% of BSA and 0.1% of NaN3 in the same 0.05 M glycin buffer. To each of the suspensions thus prepared the same amount of each of the monoclonal antibody sensitized particles was added and mixed. The mixtures were stored at 4°C.

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  • Health & Medical Sciences (AREA)
  • Life Sciences & Earth Sciences (AREA)
  • Engineering & Computer Science (AREA)
  • Immunology (AREA)
  • Hematology (AREA)
  • Molecular Biology (AREA)
  • Biomedical Technology (AREA)
  • Chemical & Material Sciences (AREA)
  • Urology & Nephrology (AREA)
  • Biotechnology (AREA)
  • Microbiology (AREA)
  • Cell Biology (AREA)
  • Food Science & Technology (AREA)
  • Medicinal Chemistry (AREA)
  • Physics & Mathematics (AREA)
  • Analytical Chemistry (AREA)
  • Biochemistry (AREA)
  • General Health & Medical Sciences (AREA)
  • General Physics & Mathematics (AREA)
  • Pathology (AREA)
  • Measuring Or Testing Involving Enzymes Or Micro-Organisms (AREA)
  • Investigating Or Analysing Biological Materials (AREA)
EP19880119491 1987-12-03 1988-11-23 Procédé pour la détermination de facteurs de coagulation du sang et réactif utilisé dans ce but Withdrawn EP0325723A3 (fr)

Applications Claiming Priority (2)

Application Number Priority Date Filing Date Title
JP306730/87 1987-12-03
JP30673087A JPH01147367A (ja) 1987-12-03 1987-12-03 凝血因子の測定方法及び測定用試薬

Publications (2)

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EP0325723A2 true EP0325723A2 (fr) 1989-08-02
EP0325723A3 EP0325723A3 (fr) 1990-11-07

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Cited By (6)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
EP0417818A1 (fr) * 1989-09-15 1991-03-20 Curative Technologies, Inc. Sélection quantitative de sécrétions plaquettaires pour un traitement efficace des tissus
AU620799B2 (en) * 1987-03-06 1992-02-27 Dade Behring Marburg Gmbh Process for the preparation of factor viii:c-deficient plasma, and a deficient plasma obtained in this way
FR2691546A1 (fr) * 1992-05-19 1993-11-26 Leroy Olivier Traceurs particulaires directs, utilisation pour la mesure et le dosage d'anticorps et d'antigène.
US5599558A (en) * 1989-09-15 1997-02-04 Curative Technologies, Inc. Selecting amounts of platelet releasate for efficacious treatment of tissue
WO2002090989A2 (fr) * 2001-05-10 2002-11-14 Holger Kiesewetter Procede permettant de detecter des antigenes de globules sanguins et des anticorps diriges contre ces antigenes
CN104198724A (zh) * 2014-08-14 2014-12-10 上海睿康生物科技有限公司 纤维蛋白或纤维蛋白原降解产物检测试剂盒

Families Citing this family (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
JP4731338B2 (ja) * 2006-02-02 2011-07-20 株式会社日立製作所 エレベータ誘導装置

Citations (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
EP0074271A1 (fr) * 1981-09-08 1983-03-16 Ortho Diagnostic Systems Inc. Liaison de deux anticorps

Family Cites Families (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
JPS6093353A (ja) * 1983-10-27 1985-05-25 Wako Pure Chem Ind Ltd Ｆｄｐの免疫学的測定方法
JPS60233553A (ja) * 1984-05-04 1985-11-20 Dai Ichi Pure Chem Co Ltd フイブリノゲン・フイブリン分解産物の測定方法

Patent Citations (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
EP0074271A1 (fr) * 1981-09-08 1983-03-16 Ortho Diagnostic Systems Inc. Liaison de deux anticorps

Non-Patent Citations (3)

* Cited by examiner, † Cited by third party
Title
CHEMICAL ABSTRACTS, vol. 104, no. 17, 28th April 1986, page 324, abstract no. 144820f, Columbus, Ohio, US; M.J. ELMS et al.: "Rapid detection of cross-linked fibrin degradation products in plasma using monoclonal antibody-coated latex particles", & AM. J. CLIN. PATHOL. 1986, 85(3), 360-4 *
THROMBOSIS RESEARCH, vol. 44, 1986, pages 715-728, Pergamon Press Plc, US; M. MIRSHAHI et al.: "A latex immunoassay of fibrin/fibrinogen degradation products in plasma using a monoclonal antibody" *
THROMBOSIS RESEARCH, vol. 50, 1988, pages 469-479, Pergamon Press Plc, US; Y. SAKAI et al.: "Determination of FDP in human plasma by a novel immunoassay" *

Cited By (8)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
AU620799B2 (en) * 1987-03-06 1992-02-27 Dade Behring Marburg Gmbh Process for the preparation of factor viii:c-deficient plasma, and a deficient plasma obtained in this way
EP0417818A1 (fr) * 1989-09-15 1991-03-20 Curative Technologies, Inc. Sélection quantitative de sécrétions plaquettaires pour un traitement efficace des tissus
FR2652088A1 (fr) * 1989-09-15 1991-03-22 Curative Tech Inc Procede de preparation d'un produit a base de substance liberee par les plaquettes, et produit obtenu par ce procede.
US5599558A (en) * 1989-09-15 1997-02-04 Curative Technologies, Inc. Selecting amounts of platelet releasate for efficacious treatment of tissue
FR2691546A1 (fr) * 1992-05-19 1993-11-26 Leroy Olivier Traceurs particulaires directs, utilisation pour la mesure et le dosage d'anticorps et d'antigène.
WO2002090989A2 (fr) * 2001-05-10 2002-11-14 Holger Kiesewetter Procede permettant de detecter des antigenes de globules sanguins et des anticorps diriges contre ces antigenes
WO2002090989A3 (fr) * 2001-05-10 2003-06-12 Holger Kiesewetter Procede permettant de detecter des antigenes de globules sanguins et des anticorps diriges contre ces antigenes
CN104198724A (zh) * 2014-08-14 2014-12-10 上海睿康生物科技有限公司 纤维蛋白或纤维蛋白原降解产物检测试剂盒

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EP0325723A3 (fr) 1990-11-07
JPH01147367A (ja) 1989-06-09

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